Plasma lipid levels are heritable risk factors that contribute to the development of coronary artery disease. Human Genome-Wide Association Studies (GWAS) have discovered more than 150 genome-wide significant loci associated with plasma lipids. A major challenge in GWAS is to identify and validate causal genes within the identified loci. Here, we developed an integrative data-driven approach that leverages publicly available data from the mouse along with existing human lipid GWAS data to identify causal genes. Through our approach, we identified Sestrin1 as an undescribed gene associated with human variation in plasma cholesterol levels. Sestrin1 belongs to a family of conserved Sestrin proteins that are known to inhibit the mammalian Target of Rapamycin Complex 1 (mTORC1) signaling pathway. Our data show that (a) liver transcript levels of Sestrin1 in the mouse reproducibly cluster with a module of genes highly enriched with cholesterol biosynthetic genes and that (b) Sestrin1 lies within a GWAS locus associated with plasma total cholesterol, in two independent human GWAS. Our experimental studies demonstrate that in cultured cells, Sestrin1 can regulate cellular cholesterol levels. Additionally, in mice, Sestrin1 global knockout mice have significantly increased plasma cholesterol levels. Collectively, our data identify Sestrin1 as a causal gene within a human GWAS locus associated with plasma cholesterol levels and establish an unrecognized role for Sestrin1 in the regulation of cellular and plasma cholesterol levels. Based on our supporting data, we propose the hypothesis that hepatic Sestrin1 regulates plasma cholesterol levels by inhibiting cholesterol biosynthesis through the mTORC1 signaling pathway. In Aim 1, we will test the hypothesis that hepatic Sestrin1 regulates plasma cholesterol levels through cholesterol biosynthesis. The tissue-specific function of Sestrin1 is unknown. Using a liver-specific Sestrin1 knockout mouse model, we will perform gain and loss of function studies to determine the role of hepatic Sestrin1 in regulating plasma cholesterol levels and cholesterol biosynthesis. In Aim 2, we will test the hypothesis that Sestrin1 interacts with the mTORC1 signaling pathway to regulate plasma cholesterol levels. It is unknown if the effect of Sestrin1 on cholesterol metabolism requires mTORC1 signaling. Using mice in which hepatic mTORC1 is constitutively inactivated or activated, we will determine if the effect of Sestrin1 on plasma cholesterol requires mTORC1 signaling. The proposal will build upon our preliminary studies and ultimately enhance our understanding of the molecular mechanisms that contribute to variation in plasma cholesterol in humans, which could lead to therapeutic strategies targeting Sestrin1 to lower plasma cholesterol levels.